Cystic fibrosis is mostly caused by the F508del mutation, which impairs CFTR protein from exiting the endoplasmic reticulum due to misfolding. VX-809 is a small molecule that rescues F508del-CFTR localization, which recently went into clinical trial but with unknown mechanism of action (MoA). Herein, we assessed if VX-809 is additive or synergistic with genetic revertants of F508del-CFTR, other correctors, and low temperature to determine its MoA. We explored and integrated those various agents in combined treatments, showing how they add to each other to identify their complementary MoA upon correction of F508del-CFTR. Our experimental and modeling data, while compatible with putative binding of VX-809 to NBD1:ICL4 interface, also indicate scope for further synergistic F508del-CFTR correction by other compounds at distinct conformational sites/cellular checkpoints, thus suggesting requirement of combined therapies to fully rescue F508del-CFTR.

The profile of PC945, a novel triazole antifungal designed for administration via inhalation, was assessed in a range of in vitro and in vivo studies. PC945 was characterized as a potent, tightly binding inhibitor of Aspergillus fumigatus sterol 14α-demethylase (CYP51A and CYP51B) activity (50% inhibitory concentrations [IC50s], 0.23 μM and 0.22 μM, respectively) with characteristic type II azole binding spectra. Against 96 clinically isolated A. fumigatus strains, the MIC values of PC945 ranged from 0.032 to >8 μg/ml, while those of voriconazole ranged from 0.064 to 4 μg/ml. Spectrophotometric analysis of the effects of PC945 against itraconazole-susceptible and -resistant A. fumigatus growth yielded IC50 (determined based on optical density [OD]) values of 0.0012 to 0.034 μg/ml, whereas voriconazole (0.019 to >1 μg/ml) was less effective than PC945. PC945 was effective against a broad spectrum of pathogenic fungi (with MICs ranging from 0.0078 to 2 μg/ml), including Aspergillus terreus, Trichophyton rubrum, Candida albicans, Candida glabrata, Candida krusei, Cryptococcus gattii, Cryptococcus neoformans, and Rhizopus oryzae (1 or 2 isolates each). In addition, when A. fumigatus hyphae or human bronchial cells were treated with PC945 and then washed, PC945 was found to be absorbed quickly into both target and nontarget cells and to produce persistent antifungal effects. Among temporarily neutropenic immunocompromised mice infected with A. fumigatus intranasally, 50% of the animals survived until day 7 when treated intranasally with PC945 at 0.56 μg/mouse, while posaconazole showed similar effects (44%) at 14 μg/mouse. This profile affirms that topical treatment with PC945 should provide potent antifungal activity in the lung.

The antifungal effects of the novel triazole PC1244, designed for topical or inhaled administration, against Aspergillus fumigatus were tested in a range of in vitro and in vivo studies. PC1244 demonstrated potent antifungal activities against clinical A. fumigatus isolates (n = 96) with a MIC range of 0.016 to 0.25 μg/ml, whereas the MIC range for voriconazole was 0.25 to 0.5 μg/ml. PC1244 was a strong tight-binding inhibitor of recombinant A. fumigatus CYP51A and CYP51B (sterol 14α-demethylase) enzymes and strongly inhibited ergosterol synthesis in A. fumigatus with a 50% inhibitory concentration of 8 nM. PC1244 was effective against a broad spectrum of pathogenic fungi (MIC range, <0.0078 to 2 μg/ml), especially Aspergillus terreus, Trichophyton rubrum, Candida albicans, Candida glabrata, Candida krusei, Cryptococcus gattii, Cryptococcus neoformans, and Rhizopus oryzae. PC1244 also proved to be quickly absorbed into both A. fumigatus hyphae and bronchial epithelial cells, producing persistent antifungal effects. In addition, PC1244 showed fungicidal activity (minimum fungicidal concentration, 2 μg/ml) which indicated that it was 8-fold more potent than voriconazole. In vivo, once-daily intranasal administration of PC1244 (3.2 to 80 μg/ml) to temporarily neutropenic, immunocompromised mice 24 h after inoculation with itraconazole-susceptible A. fumigatus substantially reduced the fungal load in the lung, the galactomannan concentration in serum, and circulating inflammatory cytokine levels. Furthermore, 7 days of extended prophylaxis with PC1244 showed in vivo effects superior to those of 1 day of prophylactic treatment, suggesting accumulation of the effects of PC1244. Thus, PC1244 has the potential to be a novel therapy for the treatment of A. fumigatus infection in the lungs of humans.

The discovery of a novel series of therapeutic agents that has been designed and optimized for treating chronic obstructive pulmonary disease is reported. The pharmacological strategy was based on the identification of compounds that inhibit a defined subset of kinase enzymes modulating inflammatory processes that would be effective against steroid refractory disease and exhibit a sustained duration of action after inhaled delivery.

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